Development and use of field application vectors to express nonadaptive foreign genes in competitive environments

Many potential applications of genetically engineered microorganisms in environmental and agricultural biotechnology involve introducing genetic capabilities into nonsterile competitive environments in which they provide no advantage to the host. Field application vectors have been designed for the purpose of creating a temporary niche for the host in such environments. This technique involves the addition to the target environment of a selective substrate readily utilizable by the host microorganism but unavailable to most indigenous species. Thirteen nonionic and anionic detergents, representing a wide range of structural complexities and molecular weights, were screened as potential selective substrates. Competition experiments in soil, using Warburg respirometry, indicated that isolates from six different detergent enrichment cultures were more active on their corresponding detergents than the indigenous microorganisms. Detergents of intermediate structural complexities and molecular weights were most effective for use as selective substrates. A field application vector that utilizes 1.0% Igepal CO-720 (detergent) as the selective substrate and Pseudomonas paucimobilis 1IGP4 as the host was tested for its ability to increase the presence of nonadaptive tetracycline resistance marker genes in soil. In soil amended with the selective substrate, strain 1IGP4 plate counts increased by three orders of magnitude and tetracycline-resistant transformant (pRK293) counts increased from 1.8 x 10/g of soil to 4.3 x 10/g in 2 days. Inoculation in the absence of substrate amendment or amendment with a nonselective substrate did not result in growth of strain 1IGP4. These results demonstrate the effectiveness of field application vectors for increasing the concentration of nonadaptive genes in competitive environments.